Table 2 Possible mechanisms underlying the five largest factors affecting DHFR abundance
EffectCategoryMagnitudeMechanistic interpretation
Embedded Image: A26T: L28RSpecies × SNP × SNP (third-order)+1.59The strongly positive effect of this third-order interaction is emblematic of the restorative effects of A26T:L28R, even on backgrounds typified by low availability, as in L. grayi (effect size = −0.84).
Embedded ImageSpecies (main effect)−1.01As described in Table 1 (as applied to its effect on Embedded Image), the C. muridarum amino acid background has low functional availability and low catalytic efficiency. These factors contribute to its negative impact on both Embedded Image and abundance.
Embedded Image:L28RSpecies × SNP (second-order)−1.01The L28R mutation, in isolation, is associated with DHFR thermostability and, relatedly, abundance (effect size = 0.88). The L. grayi background has a net negative effect on abundance (effect size = −0.84). Therefore, one might predict that their combination might cancel out toward a nearly neutral effect. Instead, this interaction has a net negative effect on abundance, an example of how some effects cannot be easily interpreted from knowledge of the underlying biochemistry of the enzyme.
L28RSNP (main effect)+0.88The L28R SNP has a strong positive effect on DHFR thermostability, which is at least partly correlated with protein abundance.
Embedded Image: GroEL+: A26T: L28RSpecies × PQC × SNP × SNP (fourth-order)+0.87The interaction between the A26T:L28R double mutant and the L. grayi amino acid background has a strongly positive effect on abundance (effect size = 1.59) that is somehow diminished in the presence of the GroEL+ PQC background. This is peculiar when we consider the positive GroEL+ main effect (effect size = 0.25). This implies that the positive effect (in terms of magnitude and direction) of the GroEL+ PQC background is specific to the SNP and amino acid combinations present in DHFR, a finding for which there is no simple, intuitive explanation.
  • DHFR, dihydrofolate reductase.